Stable isotope methodologies (mainly 13C nmr and 2H nmr) will be used to investigate several reactions involved in he metabolism of short-chain fatty acids or branched-chain amino acids. The stereochemistry of hydration of Beta-methylcrotonyl CoA will be investigated using purified enoyl CoA hydratase (crotonase) in an attempt to determine why Beta-methylcrotonate hydrates non-stereospecifically in biotin-deficient rats. The stereochemistry of isomerization of Beta-methylvinylacetyl CoA to Beta-methylcrotonyl CoA will be determined, as a model for the analogous isomerization occurring in the metabolism of unsaturated fatty acids. The stereochemicstry of dehydrogenation of isovaleryl CoA by purified isovaleryl CoA dehydrogenase will be determined. The stereochemistry of the biosynthesis of D-pantoate from deuterated formate by ketopantoate hydroxymethyltransferase, a methylenetetrahydrofolate-dependent enzyme, will be examined. The stereochemistry of the oxidative decarboxylation of Beta,Beta-dimethyl-D-malate in the degradation of pantoate will be determined. The stereochemistry of ring expansion in the conversion of penicillins to cephalosporin C will be determined, using a chiral-methyl substrate, and with the application of tritium nmr spectroscopy. Also the stereochemistry of the primary hydroxylation reaction leading from deacetoxycephalosporin C to deacetylcephalosporin C will be determined.